With the aid of protein kinase and a phosphatase purified from brain, we have studied the ability of MAP-2 (the principle microtubule-associated protein of brain) to promote microtubule assembly, when prepared in defined states of phosphorylation. With high-phosphate MAP-2 (27 mols/mol) assembly rate was decreased, average length and critical concentration were unchanged, and rates of subunit loss were increased at both microtubule ends. Only half of the MAP-2 phosphates were released by the brain phosphatase - these sites appear relatively concentrated in the binding domain, or at its junction with the projection domain. We have also found that effects of ATP on assembly cannot be entirely attributed to MAP-2 phosphorylation. Fifty percent inhibition was observed without (less than 0.1 mol/mol) protein phosphorylation; in the presence of kinase and other MAPs ATP had variable, profound and as yet enigmatic effects. For our second project, tubulin tyrosinolation, we have established facilities for HeLa cell culture, to reinvestigate initially the apparent inability to chase out tyrosine added post-translationally in vivo. Pre-column dabsylation followed by reverse phase hplc provided a rapid method to analyze changes in the concentration and specific radioactivity of intracellular free tyrosine. It appears now that tyrosine can in fact be chased out of tubulin if the free tyrosine is sufficiently diluted.